Project Summary This project will culminate in the development of a combinatorial therapy that enhances high-frequency irreversible electroporation (H-FIRE) focal ablation, surpassing traditional therapies in terms of ability to selectively target infiltrative cells beyond the tumor margin of glioblastoma (GBM). H-FIRE is a new, minimally invasive ablation technique that involves delivering a series of electric pulses that are low in energy, but intense (~1000 V) and short (~1 us) to targeted tissue for approximately 5 minutes. These pulses destabilize the cell membranes of the targeted tissue, inducing cell death without causing thermal damage. H-FIRE creates complete and predictable cell ablation with a sharp transition between normal and necrotic tissue. Furthermore, H-FIRE preserves important tissue components such as extracellular matrix, myelin sheaths, blood vessels, connective tissue, and nerves. We hypothesize that infiltrative cells (beyond the H-FIRE treated zone) can be selectively killed using a low dose of an anti-GBM drug in combination with H-FIRE, resulting in complete regression of tumors while preventing infiltration beyond the tumor margins. For tumor cells outside the zone of tissue ablation, there is a non-destructive increase in blood-brain barrier permeability, thus, making them more susceptible to the administered agents and thus making the combination of IRE and adjuvant agents synergistic. By focusing on brain cancer, we will be directly addressing the need to develop alternative approaches to radiation and chemotherapy, both of which have adverse side effects and limited efficacy. The project has three Specific Aims. In Aim 1, we will develop optimized treatment parameters for H-FIRE targeting penetration into the infiltrative niche of GBM, with a combination of H-FIRE and delivery of liposomal doxorubicin tested in a 3D micro-engineered tumor/blood-brain-barrier model (BBB). In Aim 2, we will leverage rodent models of invasive GBM for both 3D model validation, and testing of the efficacy of combinatorial treatment protocols in a more physiological relevant in vivo setting. In Aim 3, we will assess our combinatorial treatment strategy to treat spontaneous brain tumors in canine patients. If successful, this study will provide the foundation for a new form of cancer therapy capable of surpassing conventional treatments for targeting of the bulk tumor, as well as the infiltrative GBM cells beyond the tumor margin. If successful, this hybrid approach will eliminate the likelihood of tumor recurrence, while preserving the vital healthy surrounding tissue and minimizing the adverse side effects that are associated with standard therapies.